Alkyl mercurials have emerged during the last few decades as widespread environmental contaminants and threat to public health. In this proposal we will analyse the sequential biochemical and early ultrastructural events induced by methyl mercury (MeHg) in cerebellar neuronal culture and hepatocyte culture. Previous studies identified in vivo and in vitro MeHg inhibition of brain aminoacyl t-RNA synthetases and inhibition of transcription associated with impaired phosphorylation of uridine nucleotides. Recent data reveals that MeHg-induced changes in ATP reduction or diminished macromolecular synthesis are not causal to the rapid onset of granule cell death, implying an alternate mechanism to account for the lethal change, hypothesized to be due to lipoperoxidation injury secondary to free radical formation and peroxidative stress. The effect of MeHg on lipid peroxidation, glutathione content, glutathione peroxidase and free radical formation will be examined. The effect of modulating glutathione content, intracellular Ca2+ or pathways of peroxide dismutation in culture prior to MeHg addition will be examined. The dose-response and evolution of these neurotoxic events will be compared in a non-neuronal e.g. hepatocyte culture. The effect of MeHg on protein phosphorylation in culture will be determined with special reference to the cytoskeletal proteins tau, tubulin and microtubule accessory proteins (MAPs). Biochemical studies will be correlated with tubulin immunoperoxidase histochemistry, and quantitative ultrastructural morphology of microtubule configuration and ribosome-membrane interaction. Scanning electron microscopy will be used to examine changes in surface membrane morphology and provide semi-quantitative data of surface bleb formation as manifestation of early cytotoxicity. Cell culture studies allow for comparison with non-neuronal systems and provide paradigms for sophisticated metabolic labelling studies. The relative contributions of peroxidative stress, glutathione metabolism, free radical injury, and intracellular (Ca2+) will be determined. The long term goal from such studies is the development of rational therapeutic protocols and greater understanding of the biochemical pathways leading to organo-metal lethal cell injury.